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Dephosphorylation of biomolecules under the catalysis of alkaline phosphatase (ALP) is a critical physiological process. Abnormal levels of ALP activity have been associated with a number of diseases; thus, a simple and sensitive assay of ALP activity is highly demanded. Herein, to simulate biological conditions, we labeled a hydrosoluble phosphorylated heptapeptide Gly-Pro-Gly-Asn-p-Tyr-Gly-Ala (pGA) with aminated heptamethine cyanine dye (Cy) to give a low fluorescent labeled peptide Cy-pGA. The synthesized Cy-pGA and Eu3+-doped oxide Y0.6Eu0.4VO4 nanoparticles (NPs) were employed respectively as acceptor and donor to in situ form a non-fluorescent Fluorescence Resonance Energy Transfer (FRET) Cy-pGA-NP system, with the help of the strong interaction between Eu3+ ions in the NPs and phosphate group in Cy-pGA. The breaking of the FRET system of Cy-pGA-NP was triggered by the removal of phosphate group in Cy-pGA catalyzed by ALP and resulting in the release of fluorescent Y0.6Eu0.4VO4 NPs. Thus, the formed Cy-pGA-NP as a sensitive sensor can very well respond to the activity of ALP by measuring the time-resolved fluorescent intensity at near-infrared 617 nm (λ ex = 320 nm, delay time 400 μs). This sensor can not only accurately measure the activity of ALP (1–5 mU/mL) in the designed solutions, but it can also be applied to detect the activity of ALP in biological samples, such as cell lysate and human serum, without the interference of autofluorescent background of biosamples and screen ALP inhibitor by a simple mix-and-measure manner.

A non-fluorescent self-assemblage of Y 0.6 Eu 0.4 VO 4 nanoparticle (NP) and heptamethine cyanine dye bearing phosphate group p-Cy7-A, NP-p-Cy7-A, was the first constructed and used as an alkaline phosphatase (ALP) sensor. Herein, Y 0.6 Eu 0.4 VO 4 NP and new synthesized p-Cy7-A were employed respectively as donor and acceptor of Fluorescence Resonance Energy Transfer. With the help of phosphate group, p-Cy7-A as a dark quencher was captured on Y 0.6 Eu 0.4 VO 4 NP surface to form non-fluorescent NP-p-Cy7-A in situ. The disintegration of NP-p-Cy7-A was triggered by the removal of phosphate group in p-Cy7-A catalyzed by ALP, and released fluorescent Y 0.6 Eu 0.4 VO 4 NP. Thus NP-p-Cy7-A assemblage as a selective and sensitive sensor can very well responded to the amount and activity of ALP by measuring the fluorescent intensity at 620 nm (excited at 270 nm). This sensor can not only accurately measure the concentration of ALP (0.03–1.65 U/mL), but also be applied to detect the activity of ALP and screen its inhibitor by a simple mix-and-measure manner.

Dephosphorylation of biomolecules under the catalysis of alkaline phosphatase (ALP) is a critical physiological process. Abnormal levels of ALP activity have been associated with a number of diseases; thus, a simple and sensitive assay of ALP activity is highly demanded. Herein, to simulate biological conditions, we labeled a hydrosoluble phosphorylated heptapeptide Gly-Pro-Gly-Asn-p-Tyr-Gly-Ala (pGA) with aminated heptamethine cyanine dye (Cy) to give a low fluorescent labeled peptide Cy-pGA. The synthesized Cy-pGA and Eu